Ultrastable combined atomic force and total internal fluorescence microscope

Abstract

Combining atomic force microscope (AFM) with other microscopy techniques has expanded the range of potential applications for single molecule investigations dramatically. Particularly hybrid instruments with total internal reflection fluorescence (TIRF) excitation have opened new routes in life sciences. Here we present a novel design for such a hybrid microscope, which overcomes the limitations of conventional combinations caused by their limited mechanical stability. A thorough analysis of the noise spectra and a comparison of the different designs and the different operation modes are given. With this instrument we demonstrate single molecule manipulation by AFM and simultaneous TIRF imaging.